2,4-diaminopteridine aldehydes



Patented Jan. 26, 1954 2,4-DIAMINOPTERIDINE ALDEHYDES Harold G. Peter-ing, Kalamazoo, Mich., assignor to The Upjohn Company, Kalamazoo, Mich., a corporation of Michigan No Drawing. Application July 22, 1950, Serial No. 175,476

6 Claims.

This invention relates to 2,4-diaminopteridine aldehydes having the formula:

wherein the aldehyde or formyl group is attached to the 6- or '7-position of the pteridine ring, aldehyde derivatives such as the oximes, semi-carbazones, thiosemicarbazones, phenylhydrazones thereof, and to a method for their preparation.

The products of this invention have been prepared and found to be brown to light tan solids which are very soluble in aqueous acids and alkalis, less soluble in water at pH 7.0 and relatively insoluble in most common organic solvents. They are stable compounds and may be handled and used in further chemical reactions with no more precautions as to their stability than is customarily taken with aldehydes. They also readily yield the common aldehyde derivatives. They char and decompose without melting when heated above 300 degrees centigrade and are best characterized by their distinctive ultravoilet absorption spectra in acid and alkaline solution and the ratios of their extinction coefiicients at the absorption maxima. The freedom from the isomeric aldehyde can be ascertained from the numerical value of the pure 'Y-aldehyde. When the numerical value of the 248 mu E 335 mu 2 ratio is 1.4-1.5 the product is the 6-aldehyde while a value of 1.0-1.1 denotes the 7-a1dehyde. These aldehydes are useful as intermediates in the synthesis of folic acid antagonist and are themselves useful as anti-viral agents and enzyme inhibitors.

It is an object of this invention to provide new compounds that are valuable as intermediatesin the synthesis of folic acid acid antagonists and related compounds. A further object is the provision of 2,4-diaminoformylpteridines having the formula given above, and the oxime, phenyl hydrazone, thiosemicarbazone and other aldehyde derivatives thereof. An additional object of this invention is to provide a process whereby such compounds may be prepared from 2,4-diaminopolyhydroxyalkylpteridines containing either a terminal hydroxyl or a terminal carboxyl group. A still further object of this invention is the provision of such a process whereby the oxidation is caused to occur with lead tetraacetate or periodic acid as the active oxidizing agent. Other objects of the invention will become apparent hereinafter.

It is known that certain polyhydroxyalkyl compounds, in particular carbohydrates, are oxidized in a more or less special manner by lead tetraacetate and periodic acid to yield aldehydes and carboxylic acids rather than the usual carbondioxide and carboxylic acids. However, the perculiarities of this reaction are so numerous that the art is unable to predict what would result from the oxidation of a 2,4-diaminopolyhydroxyalkylpteridine with these reagents. It has now been found that 2,4-diamino-6-polyhydroxyalkyl and 2,4-diamino-'7-polyhydroxyalkylpteridines may be oxidized to the corresponding 2,4-diamino-6- formyl and 2,4-diamino-7-formylpteridines regardless of the stero chemical configuration or the length of the polyhydroxyalkyl side-chain or of the substituent on the terminal carbon atom in the starting compound. The 2,4-diamono-6- (and 7-) polyhydroxyalkylpteridines which are useful as starting materials in the process of this invention, and a method for their preparation, have been described and claimed in the copending applications Serial No. 175,477 and Serial No. 175,478, now Patent No. 2,568,462, filed of even date herewith. According to the method of the copending application Serial No. 175,477 2,4-diamino- 6-polyhydroxyalkylpteridines can be prepared by reacting a polyhydroxyaldehyde or a polyhydroxyketone with 2,4,5,6-tetraaminopyrimidine in aqueous solution at a pH between about 5.0 and 6.5 preferably in the presence of hydrazine. The above reaction is accelerated by moderate heating. as on a steam bath. The desired product can be precipitated from the reaction mixture by cooling and separating, as by filtration, and can then be purified by dissolving in dilute alkali, decolorizing with adsorbent charcoal and recovering by acidification and filtration or by simple washing of the original precipitate with water andsorganic solvents: The 2;4-d-iamino-'7-polyhydroxyalkylpteridines can be prepared by reacting the osone of a monosaccharide having at least four carbon atoms with 2,4,5,6-tetraaminopyrimidine-in' aquee ous solution at a pH of between about 4.5 and 6.5. The isolation of the product is'scarriedout ir'r-th'e same manner as described for tiled-isomer above; According to copending application 175,478, now Patent No, 2,568,462 using thesamerprocedure; 2,4,5,6-tetraaminopyrimidine can be condensed: with 5-ketog1uconic acid preferably in thepresence of hydrazine whereby a fi-polyhydroxyalkyl pteridine is obtained having a carboxyl group upon: the. terminal carbon atomr The 2';4-' diamino-6 --(or 7"--) polyh-ydroxyalkylpteridines thus obtanied can he -oxidized" to the 2;"4"-diamino-6= (or =7) formylpterid-inesof this -in-- venti'on by mixing'an excess of purified lead tetraacetateidissolved in warm glacial acetic acidj or a mixture: offglacialfacetic acid and acetic anhydride; with" an aqueous solution :of the p'olyhydroxyalkylpteridine. The reaction israpid andexothermic. While the. starting polyhydroxya-lkylpteridines may be di'ssolved in water-as above,

in which? they-are soluble to the amount of'about oneehalffpercent at 100? degrees centigrade,- it: is

preferred "that smalliamounts of alkali such as an'alkali' metal hydroxide, carbonate and: 'bicar bonate'or ammonium hydroxide b'e -used 'to effect solutionr. Alternatively-ythe aqueous sol'ution can be acidified to about pH3.0 prior'tothe addition of the lead: tetraacetater solution. However the use ofzsmall 'quantitiesof alkali is a preferred proceduresasfthevolume;of solution required is sufficifently reduced to be of' practical importance.

An alkaline solution can be used to dissolvethestarting .polyhydroxyalkylpteridinesince the'large excess cf acid: added;v alongwith the lead acetate assures. that the: resulting aldehyde: will "not' be destroyed: by the added alkali;

The. exothermic: nature of the reaction .causes the-temperature to rise somewhat; the final temperature being-dependent upon th'e volume of the solutions. After mixing, and' allowing-the reaction toriproceedg'withthe normal risein-temperature= causedby -the-nature of 'thereaction; the reaction: mixture =i's--b'rought--toabout- 90 to 100 degrees centigradefor -an'-- additional length of time-to completeth'e oxidatione' This a dditional time required varies; depending upon the quantity of reactants; with up-"to about one-half hour being suflicient;

Aftertheoxidation is complete; the PbO'z; formed as a byproduct of the-reaction; is-- removedpreferably by fiitratienz Thedead-acetate formed'is converted-"toinsoluble-lead sulfate by the addition of dilutesul fur-ic acid' or a sulfate salt 'suchas ammoniumsu'lfate, whereupon the lead sulfate which-precipitates is removed- After removal of the-Iead-upon cooling, the-filtrate deposit's crystals of the desired aldehyde whichare collected for-furtherpurification as desired.

The--- 2;4-diamino-6'(or 7-) polyhydroxyalkylpteridines'can al'sobe oxidizedto 2,4-diamino: 64 01" 'F-lformyl pteridineaby mixing; a solution of the 2,'4'-diaminopolyhydroxyalkylpteridine in an aqueous acid solution such as aqueous acetic or sulfuric a'ci rl with-"an aqueous selution of per iodi'c acid:- Thereaction "isquite rapidat term peratu-res "between about' and Y degrees een tigrade although temperatures of up to about degrees centigrade are satisfactory. The reaction is not exothermic so external heat must be applied when the reaction is carried out above room temperature. Aiter completion ofithe reaction theacidity of the-solutionis reduced to between about pH 4.0 to pH 6.0 by the addition of alkali the cooled reaction mixture deposit the 2,4-diamino-640cm) formyl pteridine as a solid, which is collected for use or further purification as desired:

The-following examples are given to illustrate certain" preferred embodiments of this invention but suclr modifications as are apparent to those skilled'ain the art to which this invention pertains can be made without departing from the spirit andscope of this invention.

PREPARATION. 1.2,l-DIAMINQ-6;TETRAHYnRoxYBumYL- L-XYLQ) mmerse A dry mixture was. prepared containing 1.065 grams of 2,4,5,fi-tetraaminopyrimidine dihydrochloride; 0.85 gram-of sodium bicarbonate and 1.8 grams of L-sorb'oses To the dry mixture 13 milliliters of water was added, followed by 1.0 milliliter of acetic acid and 0.6 milliliter of an 3.5spercent solution of hydrazine hydrate: The pH oftthe solutionwas-1adiustedafrorn EbOUfiPT-Q tofahout pH 5-8 by-theaddi-tionpot.glacialraceticacid; one-half, milliliter; being sufiicient; The: reaction mixture was: heated. '-ont a- Water; bath. to 95-100 deg-recs centigrade for about two'hours and-thencooled to about five: degrees; centigrade ior. 16 hours; The brown precipitate: which had' formed .was' collected, washeda-with. alcohol, acetene ether and *dried, 'I'hereewas thus; obtained 0.6 gram oil, 2,a diamino-fietetrahydroxybutyle (Lexyloi-pteridineasa. brownpcwder which had an Ina condensation-carried out 'as-above; the addition 0.60 gram ofboric' acidaft'er'completion of the heating--pe1-iod"gave-1551 grams of pteridine "as the diboric acid complex-which had an E370'mu When carried out in a like-mannerthee-cone densation of L-xylose: and .2,4,5,6-tetraaminopyrimidine gives 2A diamino6-trihydroxypropyl pteridine.

(L-XYLO -PTERIDINK To a mixture of BOOmilligrams of 2,4,5,6-tetraaminopyrimidine hydrosulfate and 600 milligrams of sodium. .bicarhonate,.-;ten milliliters of..- water added and the-mixture :stirred: vigorously The pH of the solution wasadjusted to a pH of 6.5 by the addition of sodium i acetate and then added to 32 milliliters ofa solution containing 650; milligrams of L-serbosone;=.-( also--. called; L: guiosone); One grampfiboric acid was:;added and the solutionwas;,warmedion a steam: bath until all of the reactants-were in solution after which the reaction mixture was heated to about degrees centigrade for ten minutes. The reactiommixture.wa tcooiediand the resulting precipitate was; collected and-"washed successively with 40; milliliters ofi Wat-er; ethanoland eth'erafter which it was dried. There was'thus ob tained. 690 milligrams-10f;-ZAPdiaminQJ-tetrahydroxybuty1(Lxylo)-pteridme as a tan powder whose ultraviolet absorption spectra in 0.1 normalNaOH solution showed'minima at 235 mu and 310 mu and maxima at 255 mu and 365 mu, having an ratio of 2.5. In 0.1 normal HCl maxima were observed at 240 mu, 285 mu and 335 mu and PREPARATION 3.2,4-D1AMIN0-6-0MEGA CAnBoXY- TBIHYDROXYPROPYL-PTERIDINE To a dry mixture of 1.065 grams of 2,4,5,6-tetraaminopyrimidine hydrochloride, 1.36 grams of sodium acetate trihydrate, 2.36 grams off-calcium 5-keto gluconate and 600 milligrams of boric acid a solution of 1.4 milliliters of glacial acetic acid, 0.6 milliliter of 85 percent hydrazine hydrate and milliliters of Water was added. The resulting bright yellow solution had a pH of 4.5 to 5.0. The reaction mixture was warmed to 85-95 degrees centrigrade for about 45 minutes, the solution becoming dark brown. Upon cooling a precipitate formed which was collected, washed twice with water, once with alcohol and once with ether. After drying at 60 degrees centigrade under reduced pressure there was obtained 1.74 grams of 2,4-diamino-6-omega carboxytrihy droxypropylpteridine as alight brown powder having ultraviolet absorption maxima in 0.1 normal NaOH solution at 257 mu and- 370 mu, minima at 238 mu and 322 mu and an WO mu ratio of 3.1.

One and forty one-,hundredths grams of 2,4- diamino-fi-tetrahydroxybutylpteridine was dissolved in '75 milliliters of percent aqueous acetic acid. The pH of the resulting solution was 310. To this solution 10.3 grams of PbsO4 was added portionwise with stirring over a period of thirty minutes. The reaction mixture was allowed to stand for about twenty minute after which time all of the Pb304 had gone into solution and the temperature of the reaction mixture had risen to about degrees centigrade. Two hundred milligrams of activated carbon was added to the cloudy solution and after stirring was allowed to stand for fifteen minutes, then filtered and to the clear filtrate there was added a'solution of 6.0 grams of ammonium sulfate in fifteen milliliters of Water. A white precipitate of lead sulfate appeared which wa removed and the resulting clear yellows olution divided for isolation into two portions.

Portion 1 was extracted twice with 250 milliliter quantities of ether. A yellow precipitate appeared in the aqueous layer and upon cooling further precipitation took place. The precipitate was collected, washed with alcohol then a mixture of acetone and ether, whereupon '70 milligrams of yellow 2,4-diamino-S-formylpteridine was obtained.

Portion 2 was adjusted to pH 5.0 with sodium carbonate whereupon a yellow precipitate appeared which after cooling the solution wascollected and washed with water, alcohol, ether and dried. There was thus obtained milligrams of yellow to tan 2,4-diamino-fi-formylpteridine.

The mother liquor from the two portions. which still contain aldehyde, were combined and to the solution there was added an excess of phenylhydrazine. The reaction mixture :was warmed on a steam bath to about'ninety degrees centigrade whereupon a deep red precipitate appeared which was collected after cooling the reaction mixture. The precipitate was washed with water, alcohol, and then ether and'after drying 180 milligrams of 2,4-diamino-6-formyl pteridine phenylhydrazone was obtained as a red powder.

Example 2.2,4diamino-6-formylpteridine by lead tetraacetate oxidation 1 Seven hundred milligrams of 2,4-diamino-6 tetrahydroxybutylpteridine was dissolved in fifty milliliters of tenth normal sodium hydroxide solution and warmed to about forty degrees centigrade. A solution of four grams of lead tetraacetate which had been dissolved in'25 milliliters of acetic acid and five milliliters of acetic anhydride and warmed to fifty to sixty degrees centigrade was added rapidly to the pteridine solution. A brown precipitate of lead dioxide appeared after which 1.5 grams of ammonium sulfate was added with stirring, 200 milligrams of activated carbon was added to the reaction mixture-and the solution then filtered. The orange to red filtrate was extracted four times with ether,'a brown precipitate appearing after the third extraction. The solution was then cooled and the precipitate which had formed was collected and washed with water, acetone, and ether. There was thus obtained 160 milligrams of 2,4ediamino- 6-formylpteridine as a tan powder whose ultraviolet absorption spectra in tenth normal sodium hydroxide solution showed maxima at 262 mu and 370 mu and a minimum at 315 mu with an 370 mu ratio of 2.19. In another similar reaction a 62 percent yield of aldehyde was obtained having an 352m j 370 mu ratio of 2.0. Example 3.--2,4dia1aino-6-formylpteridine thicsemicarbazone To a solution of forty milligrams of 2-,4-diamino-G-formylpteridine there was added an excess of thiosemicarbazide and the reaction mixture warmed on a steam bath for about twenty minutes. A yellow precipitate formed, which after cooling the reaction mixture was collected, Washed with water, acetone, ether and then dried. There was thus obtained 37 milligrams of 2,4-diamino 6 formylpteridine thiosemicarbazone whose ultraviolet absorption spectra showed maxima at 265 mu, 340 mu and 405 mu and minima at 245 mu, 305 mu and 370 mu.

Example 4.2,4-' diamino 6 formylpteridine oxime v To a solution of 500 milligrams of 2,4-diaminofi-formylpteridine in a mixture of water and acetic acid, an aqueous solution containing milligrams hydroxylamine hydrochloride was added and the pH of the resulting deep yellow solution adjusted to between 6 and '7 by the addition of sodiumzhydmxidessoiutiomi 'ILnesprecipi-tatezwhich formeckfwasicollectedcawashedrwitmwater,-acetone; etherrandidried; Thereawas's'thusrobtained fifty li rams: of: 2;4 diaminoefi=formylpteridineoximezas aatan powder:-whose ultraviOletzabsorption spectra: im;0.08-;= normal sodium hydroxide solutiomshowed.zmaxima: 2311262 mu and: 382' :mu with at minimum; at; 342 Inn and 24o-mu and: a" point of infiectioniatiaboutz3o5zmu;,

Ett'umpie" 5 i--2 ;4'- diaminoi- 6-formylpteridine Seven hundred; and ninety millig-rams of: 2,4: diamino .--6-12-;22.,3t-trihydroxyi -carboxypropyle pteridi-nee. -.2ag-ra-ms 10f sodium acetate and i100 milliliters-oi watenwere .warmed on. :a steam-bath; Ammonium hydroxide was added to theawarm solution whereupon a. dark brown solution of the pteridine' was obtained: While 'maintaining' the reaction temperature" between about fifty and about: sixty degrees: centigrade, ansolution: of 2.6 grams: of. lead. tetraacetatea; dissolved in. a; mix-; ture: of; seventeen: milliliters 1' of acetic acid; ten millilitersof. acetic :anhydride-andTfii ty milliliters of.- absolute methanol was ia'ddedstof the .:pteridine solution; The 2' reaction temperature? was main tained at about fif-ty to; about sixty. degrees centigra'detforran additional twenty minuteszwher'er upon: it --.wasv broughtto boiling and 1 filtered f to removeethe lead fdioxide which .had precipitated. The reaction'zmixturewvas: then: cooled: and-slight excesssof diluteasulfuric acid added'zandithe lead sulfatez -whichiformed was removed: Upon-:furthey cooling. the-brownrprecipitatei which ape neared. was collected; and: after? washing: with water; alcoh'ob. ether:v arrdzdrying 200.:milligrams off. 2Adiamino-6-+formylpteridine was obtained asra brownapowder:

Efiiample "6;"2,'4-=dz'ami1to'- 7 formylpterz'dine by perioddte and lead'tetmacetatet'omidation' Two; hundred r and five; milligrams: of, 2,14=die amino '7 .-tetrahydroxybutyl (L-xylo) -pteridine was. suspended in 1 five: milliliters ::of 1 water: and sulfuric acid added untiLaclear solution was obtained. With stirring; 350 milligrams of HIO4.2H2O dissolved one milliliter of water was. addedv'in one: portiomz. an: orange colored solutiomresultingz; Activatedecharcoal, fiftyzmih ligrams (Darco G-60) was. added and the reaction mixture stirred at room temperature for an When 205 milligrams .of 2,4-diamino-7-tetrae hydroxybutyl (L-xy1o)..-pteridine was oxidized with lead tetraacetate as in Example 2, fifty as milligrams: 24512diaminopteridinesqaldehydeshaare ingrthe: same ultraviolet; absorption: spectra: and. extinction coefficient ratio as that obtainedbs. the periodate oxidation.

Example 7.2,4-diamino-.6'- formylpteridine by periodate oxidation To a solution of 510milligramsof 2,4-diamin'oe- G-tetrahydroxybutyl (L-xylo). pteridine in eleven milliliters of two normal acetic acid was added 855 milligrams of HIO'4.2H2'O dissolved in two milliliters of water? After standing terl minutes at room temperature: the" precipitate which formed :was collected; washed withacetone; ether and dried.- There was': thus' obtainew 130 mill'i grams of 2,4 -diamino 6formylpteridine: as a brown solid.

ExampZe'-8-:'Periodateoxidation To a: solution. of 2.55 grams OfiHIOLfZI-Izfi) dissolvedrinth'irtyetwmmilliliters :of I'two normal suinfurieacidat: a t'emperature 0f 35" to: 40. degrees centigra'de, gram. of 2;4 -diamino?-6-'tetrahy droxybutyl-(Dl-arabo.) p'teridine' was-i add'ed'iporz tionwise; During'the addi'tionai-small amount of purple vapor; appeared. After: the addition 1 was complete the reaction mixture was' Warmedto about sixty degrees 'centigrade. Activatedchar coal; 200 milligrams, wasaddedwitli stirring-and after standing forabout fifteen 'minutes the reaction' mixture was. filtered; ThepH of theifiltrate' was adjusted. to about-.420 byithea'dditionof sodi' um carbonate and sodium acetate; 'On coolinga heavypr'ecipitate separated 'whichwasa'collected; washed three times with water; twice with alco hol and once-with ether. Aiterdrying'un'd'er re; ducedpressure at about sixty degrees :centigrade 300 milligrams of 2," diamino-6-formylptridinewas obtained as a yellow-solid."

Iclaim:

1. A compound selctedLfrom the group consisting of 2,4-diamino-6 formylpteridines, 2,4- diamino-7-formylpteridines and aldehyde deriv ativesthereof.

2. 2,4-diamino=fi-iormylpteridine.

3. 2,4-diamino-7-formylpteridine.

4'. 2,4-diamino-6-formylpteridineoxime:

5': 2,4'- di'amino 6=formylpteridinethiosemicar baz'one:

6i 214=diam-ino=6-formylpteridinephenylhydi'a zone.

HAROLD G; PETERING'E' References Cited" in the file? ofthis patent UNITED STATES" PATENTS Number: Name": Date 2,520 156: Lindl-ar 'et al; Aug-. 29; 1950" FOREIGN" PATENTS Number. Country. Date- 626,171 Great Britain. July. 11;..1949' OTHER REFERENCES? Skipper etalf, Cancer 3, 348-353 11950). Daniel et al., J.'Biol.'Ch'em. 169, 689-697 (19477;

Petering et -al., J. Am. Chem. Soc;, 69,12566- 2567 (1947).

Seeger et al., J. Am. Chem. so-e; 69;".2567 (1947 )Q' 

1. A COMPOUND SELECTED FROM THE GROUP CONSISTING OF 2,4-DIAMINO-6-FORMYLPTERIDINES, 2,4DIAMINO-7-FORMYLPTERIDINES AND ALDEHYDE DERIVATIVES THEREOF. 